The liver cells were incubated with Fc-blocking reagent (TrueStain, Biolegend) for 5 min, incubated with APC-SIINFEKL-H-2Kb dextramers (Immunotools) for 10 min and then stained with the appropriate antibodies detailed in the supplementary files. For the assessment of the MHC class I molecule Kb bound to the ovalbumin peptide SIINFEKL, RIL-175-LV-OVA-GFP or RIL-175-LV-GFP cells were left untreated or incubated overnight with 250 ng/ml γIFN (Peprotec) and then labeled with the APC anti-SIINFEKL-H2Kb antibody (clone 25-D1.16). The samples were processed with an Attune NxT Cytometer (ThermoScientific) and analyzed with FloJo (Treestar). Results of FACS analysis are presented as the percentage of a population or as mean fluorescence index (MFI).
Protocol detail
SIINFEKL-specific CD8+ T Cell Assay
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Anti antibody
Antibodies
Cells
Clone
Fluorescence
Liver cells
Mhc class i molecule
Ovalbumin
Siinfekl peptide
Corresponding Organization : University of Geneva
Other organizations : University Hospital of Geneva
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Variable analysis
independent variables
- Incubation with Fc-blocking reagent (TrueStain, Biolegend) for 5 min
- Incubation with APC-SIINFEKL-H-2Kb dextramers (Immunotools) for 10 min
- Incubation of RIL-175-LV-OVA-GFP or RIL-175-LV-GFP cells with 250 ng/ml γIFN (Peprotec) overnight
- Percentage of a population
- Mean fluorescence index (MFI)
- Appropriate antibodies detailed in the supplementary files
- Untreated RIL-175-LV-OVA-GFP or RIL-175-LV-GFP cells
- RIL-175-LV-OVA-GFP cells incubated with 250 ng/ml γIFN (Peprotec) overnight and labeled with the APC anti-SIINFEKL-H2Kb antibody (clone 25-D1.16)
- RIL-175-LV-GFP cells left untreated and labeled with the APC anti-SIINFEKL-H2Kb antibody (clone 25-D1.16)
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